(1) Field of the Invention
The present invention relates to a gene involved in foam formation during microorganism culture. The present invention further relates to an acetic acid bacterium which is bred by reducing or deleting the function of a protein encoded by the gene involved in foam formation and which thus has a suppressed foaming ability during culture and generates a larger amount of acetic acid. The present invention still further relates to a method for producing vinegar using the acetic acid bacterium, and vinegar produced by the production method.
(2) Description of Related Art
Foam formation during microorganism culture is a major concern in the food industry and chemical industry where microorganisms are utilized. In many cases, foam is generated when microorganisms are cultured, especially when cultured under aeration and agitation. A foam layer is formed at the upper part of the culture tank and raises problems such as reduction of the working volume in the culture tank, or loss of the culture broth, compositional change of the broth and leakage of the microorganisms due to the outflow of the foam from the upper part of the tank. As such, foam formation causes problems including decrease in the production efficiency and deterioration in quality as well as environmental problems. For this reason, it has been a critical object to suppress foam formation when obtaining products by an efficient culture of microorganisms. Decrease in the production efficiency due to foam formation has similarly been a problem in vinegar production using an acetic acid bacterium.
Therefore, physical and chemical methods have been developed as defoaming methods. Physical methods include a mechanical method in which a shear force is applied to foam by such means as an agitating blade to destroy the foam, thermal method in which a liquid viscosity is decreased by heating to destabilize the foam, and electric method in which foam is broken by means such as energization, sparking and electric current. All of these methods, however, raise cost by introducing and using equipment. In addition, defoaming results brought by these methods have been insufficient.
Chemical methods include adding antifoaming agents. Compounds such as alcohols, esters, fatty acids and silicon are used as antifoaming agents. However, while a defoaming method using an antifoaming agent is simple, there have been problems caused by some antifoaming agents such as decrease in the oxygen transfer rate which is important for microorganism growth and the material production, inhibition of the microorganism growth, and adverse influence on the isolation and purification steps.
Although the foam formation mechanism during microorganism culture remains largely unknown, some genes and proteins involved in foam formation in eukaryotes have been found. One of those genes is the awal gene, which is involved in foam formation and found in yeast (see for example, Non-patent document 1). This gene encodes the glycosylphosphatidylinositol anchor protein which is a protein specific to eukaryotes and this protein is involved in the cell surface hydrophobicity, and foam formation is suppressed when the gene encoding this protein is disrupted.
Further, a protein called Hydrophobin which is either hydrophobic or amphipathic was found in fungi, mushrooms, etc., and it has been found that foam formation is suppressed by disrupting the gene encoding Hydrophobin (see for example, Patent document 1).
In prokaryotes, however, knowledge of genes or proteins involved in foam formation during culture is scarcely obtained. For prokaryotes, therefore, breeding of bacterial strains with less foam formation has been desired as a novel defoaming means to replace the physical or chemical methods.
The presence of an intercellular signal communication system in which transcription of specific genes is controlled depending on the cell density has been recently elucidated in many bacteria. This system is called quorum-sensing system (a control system sensing cell density) and is involved in the expression control for various functions such as bioluminescence, exoenzyme production, virulence expression, biofilm formation, and antibiotic production.
For example, two kinds of proteins are involved in the quorum-sensing system which has been found in many Gram negative bacteria such as Vibrio fischeri (see for example, Non-Patent document 2: Fukushima, Jun, “Expression of pathogenicity and quorum sensing system in Pseudomonas aeruginosa,” Bioscience and Industry, Vol. 60, No. 4, pp. 219-224 (2002)). One is an Acyl Homoserine Lactone Synthase, that synthesizes acyl homoserine lactone which is an intracellular signal molecule, and the other is an Acyl Homoserine Lactone Receptor-Type Transcription Factor, that is a receptor of acyl homoserine lactone and that also functions as a transcription factor.
Acyl homoserine lactone produced by Acyl Homoserine Lactone Synthase in a bacterial cell diffuses inside and outside the bacterial cell. As the concentration of acyl homoserine lactone is increased, it forms a complex with the Acyl Homoserine Lactone Receptor-Type Transcription Factor in the bacterial cell to control the gene transcription.
The present inventor has already obtained two kinds of genes involved in the quorum-sensing system in an acetic acid bacterium, namely, a gene encoding the Acyl Homoserine Lactone Synthase (hereinafter may be referred to as orf1) and a gene encoding the Acyl Homoserine Lactone Receptor-Type Transcription Factor (hereinafter may be referred to as orf2) and has demonstrated that the quorum-sensing system in an acetic acid bacterium is involved in the acetic acid production ability (see for example, Patent document 2).
Further, despite that the nexus between the quorum-sensing system and foam formation during microorganism culture had been totally unknown, the present inventor obtained a gene involved in foam formation during culture of an acetic acid bacterium (hereinafter may be referred to as orf3) as a gene involved in the quorum-sensing system of an acetic acid bacterium, modified the gene, and demonstrated that reducing or deleting the function of a protein encoded by the gene can suppress foam formation during culture of an acetic acid bacterium and increase the ability of an acetic acid bacterium to produce acetic acid (see for example, Patent document 3).
However, the whole picture of a family of genes involved in the quorum-sensing system of an acetic acid bacterium has not yet been sufficiently elucidated and it is possible that there are genes involved in foam formation during culture of an acetic acid bacterium other than the above genes. It has thus been expected that the gene can strongly improve the acetic acid production ability.    Non-Patent Document 1: Journal of bioscience and bioengineering, Vol. 97, No. 1, pp. 14-18, 2004.    Non-Patent Document 2: Bioscience and Industry, Vol. 60, No. 4, pp. 219-224, 2002.    Patent Document 1: Published Japanese translation of PCT international publication No. 2003-507056.    Patent Document 2: Japanese Laid-Open Patent Application No. 2008-206413.    Patent Document 3: Japanese Laid-Open Patent Application No. 2008-228660.